| Summary: | 碩士 === 國立臺灣大學 === 食品科技研究所 === 92 === Numberous bioactive components in mushrooms have been demonstarted to be revelant to the antitumor, anticancer, immunoenhancement, and differentiation-inducing effect of leukemia. Researches in the present laboratory have proved that protein extracts from AA（Agrocybe aegerita） and HM（Hypsizigus marmoreus） were responsible for the growth inhibition and differentiation of U937 cells into macrophages/monocytes. As a subsequent experiment to study the cell cycle arrest mechanisms of U937 cells by protein isolates from AA and HM, protein in mushrooms were fractionated by 100 % saurated ammonium sulfate, followed by a DEAE Sepharose CL-6B chromatography, eluted by 0.1 M NaCl for AA protein (AA-0.1) and 0.2 M NaCl for HM protein (HM-0.2).
Results showed that AA-0.1 and HM-0.2 at a dosage of 25 mg/mL strongly inhibited the growth of U937 cells by 79.9 and 78.6 %, respectively. Apparent G0/G1 phase arrest was observed when cells were incubated for 24 hr with those two proteins. Extended incubation for 72 hr with AA-0.1 and HM-0.2 revealed higher (87.2 and 89.6 %) percentage of cells arrested at this phase. Of note, no apparent apoptosis was examined during the incubation. Cold extracts and ammonium sulfate precipitates of mushrooms (AA and HM) at dosages ranged from 100 - 400μg/mL and 25 - 50μg/mL, respectively, exhibited similar G0/G1 phase arrest, regardless of the difference in the incubation time.
Western blotting analysis revealed the decrease in cdk4 with the increasing incubation time of U937 cells with AA-0.1 and HM-0.2. Sharp decrease in PCNA, which facilitates the cell proliferation, after incubation with HM-0.2 for 24 hr was detected.
Other factors related to G1 phase arrest includes D type cyclins, CDK2, CDK6 and INK4 family、CIP/KIP family in CDKI as well as Rb、E2F have been planned to be tested with Western blotting to elucidate their roles in cell cycle of U937 cells arrested at G0/G1 phase by active components from dietary mushrooms.
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